Hepatitis E virus of subtype 3i in chronically infected kidney transplant recipients in southeastern France

Abstract

Hepatitis E virus (HEV) is a leading cause of waterborne acute hepatitis in developing countries. In Europe, HEV causes a zoonotic disease and is hyperendemic in southern France. Four HEV genotypes (1 to 4) have been defined, and the most used classification divides them into 24 subtypes. Autochthonous European HEV strains belong in majority to genotype 3. Subtypes 3c, 3f, and 3e are representative of the HEV diversity in France. HEV causes chronic hepatitis in solid-organ transplant recipients in Europe, and viral characteristics associated with chronicity are poorly documented. We sequenced 343-nucleotide-long HEV genomic fragments from the serum of eight chronically infected kidney transplant recipients and a near-full-length genome in one case. We identified in four patients (50%) HEV of subtype 3i, not described previously in France. If shorter genomic fragments were used in phylogenetic analyses, these HEV sequences were clustered with open reading frame 2 (ORF2) fragments labeled as subtype 3c. At least five of the eight HEV 3i sequences recovered from humans in our phylogenetic analyses were from chronically infected kidney transplant recipients. These data show that the description of the prevalence and geographical distribution of HEV subtypes may be partially inaccurate and that criteria for classification as 3i and 3c should be clarified. Extended molecular virology analyses are required to improve knowledge of HEV epidemiology and determinants of chronic HEV infection.

FIG 1

Phylogenetic tree based on a 343-nucleotide partial sequence corresponding to nucleotides 5999 to 6342 of open reading frame 2 (ORF2) of the HEV genome (GenBank accession no. AF082843). The HEV sequences obtained in our laboratory are indicated by a black frame. The 10 sequences with the highest BLAST scores (excluding those from our laboratory or with a query coverage of <98%) recovered from the NCBI GenBank nucleotide sequence database (indicated in boldface, underlined, labeled with BBH [for “best BLAST hit”], GenBank accession number, host, country, and year of sample collection or sequence submission) with the ORF2 fragment from the near-full-length genome obtained in this study have been incorporated into the phylogeny reconstruction in addition to a comprehensive set of 245 full-length genome sequences that were downloaded from the Virus Pathogen Resource database (10) (labeled, if they are clustered with sequences from the present study, with genotype and subtype, GenBank accession number, host, country, and year of sample collection or sequence submission). To allow for the better legibility of the tree, branches were collapsed when they did not directly link to sequences from the present study and when the bootstrap value at the node was >90%. Nucleotide alignments were performed using the MUSCLE software (http://www.ebi.ac.uk/Tools/msa/muscle/). The tree was constructed using the MEGA 5 software (http://www.megasoftware.net/) and the neighbor-joining method. Branches with bootstrap values were obtained from 1,000 resamplings of the data, and values greater than 50% are labeled on the tree. The EF206691 avian HEV sequence was used as an outgroup. A single sequence name is indicated for collapsed branches. A single sequence name is indicated for collapsed branches. The scale bar indicates the number of nucleotide substitutions per site. BBH, best BLAST hit; BOL, Bolivia; GER, Germany; FRA, France; Hu, Human; JAP, Japan; KYR, Kyrgyzstan; MON, Mongolia; NLD, the Netherlands; Sw, Swine; SWE, Sweden; Wb, wild boar. See also Figure SA1 in the supplemental material.

FIG 2

Phylogenetic tree of HEV full-length or near-full-length genomes. The legend of this figure is the same as that for Fig. 1, except the abbreviation “BBH” is not used in this figure and the avian HEV sequence AY535004 was used as an outgroup.